The long term goal of this project is to study, at a biochemical level, the role of hormone-receptor interactions in controlling the synthesis and secretion of adenohypophyseal hormones. Studies will be performed using clonal strains of growth hormone- and prolactin-producing rat pituitary tumor cells maintained in tissue culture (GH-cells). Thyrotropin-releasing hormone (TRH) affects hormone production in these cells. TRH binds to specific membrane receptors, and TRH regulates the number of its own receptors. Receptors for TRH will be solubilized and the affinity of soluble receptors for TRH and TRH analogs determined. Molecular properties of soluble receptors will be compared for the TRH-receptor complex, the free TRH receptor, and the receptor from "downregulated" cells. The soluble receptor will be substantially purified, in part by affinity chromatography. To elucidate the mechanism of receptor regulation, the cellular localization and rates of synthesis and degradation will be determined for TRH receptors from control cultures and "downregulated" cells. GH-cells will be synchronized and used to determine at what point in the cell cycle hormones are synthesized, TRH binds to receptors, cells are responsive to TRH, and receptor regulation occurs. The mechanism by which thyroid hormones block the effects of TRH will be investigated. Effects of L-thyroxine and L-triiodothyronine on the biological actions of TRH and TRH-receptor interactions will be measured.